Comparison of real-time fluorescent RT-PCR and conventional RT-PCR for the detection of hepatitis E virus genotypes prevalent in China
Comparison of real-time fluorescent RT-PCR and conventional RT-PCR for the detection of hepatitis E virus genotypes prevalent in China
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DOI：
10.1002/jmv.21040
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发表时间：
2007-12-01
影响因子：
12.7
通讯作者：
Wang, Youchun
中科院分区：
文献类型：
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作者：
Zhao, Chenyan;Li, Zhuo;Wang, Youchun
为了比较实时荧光RT-PCR与常规RT-PCR的特异性和敏感性，用两种方法检测了110名健康献血员、120名临床诊断为慢性B型肝炎患者和416名非甲-丙型急性肝炎患者的血清以及HEV基因型1和4的系列稀释液。所有来自健康献血员和慢性B型肝炎患者的样本均为阴性。实时RT-PCR可检测与常规RT-PCR相同的基因型1的最终稀释度，但可检测比常规RT-PCR低10倍的基因型4的浓度。在416例临床诊断为非甲-丙型急性肝炎的患者中，实时荧光定量PCR和常规RT-PCR分别检测出127例（30.5%）和83例（20.0%）HEV阳性。实时荧光定量RT-PCR与常规RT-PCR的符合率为80.8%。171份标本中，实时荧光定量RT-PCR检测抗-HEVIgM阳性96份，常规RT-PCR检测抗-HEVIgM阳性57份; 245份标本中，实时荧光定量RT-PCR检测抗-HEVIgM阴性31份，常规RT-PCR检测抗-HEVIgM阳性26份。对常规RT-PCR阳性的所有扩增子进行测序。在83株分离株中，基因型1型7株，基因型4型76株。因此，这两种检测方法都具有很高的特异性，但实时RT-PCR检测比常规RT-PCR更敏感。此外，戊型肝炎病毒基因型4是负责大多数散发病例的戊型肝炎在中国北方。
To compare the specificity and sensitivity of a real-time fluorescent RT-PCR assay with conventional RT-PCR, sera from 110 healthy blood donors, 120 patients with a clinical diagnosis of chronic hepatitis B, and 416 patients with non-A-C acute hepatitis, as well as serial dilutions of HEV genotypes 1 and 4, were tested with both assays. All samples from healthy blood donors and patients with chronic hepatitis B were negative by both assays. Real-time RT-PCR could detect the same final dilution of genotype 1 as conventional RT-PCR but could detect a 10-fold lower concentration of genotype 4 than conventional RT-PCR. Of 416 samples from patients with a clinical diagnosis of non-A-C acute hepatitis, 127 (30.5%) and 83 (20.0%) were positive for HEV by real-time and conventional RT-PCR, respectively. The concordance of real-time and conventional RT-PCR was 80.8%. Furthermore, 96 and 57 of 171 samples were positive for anti-HEV IgM by real-time and conventional RT-PCR, respectively, and 31 and 26 of 245 samples negative for anti-HEV IgM, were positive by realtime and conventional RT-PCR, respectively. All amplicons positive by conventional RT-PCR were sequenced. Of 83 isolates, 7 and 76 belonged to genotypes 1 and 4, respectively. Thus, both assays have a high specificity, but the real-time RT-PCR assay is more sensitive than conventional RT-PCR. Furthermore, HEV genotype 4 is responsible for most sporadic cases of hepatitis E in the north of China.